The present invention relates to carbohydrate mimetics that inhibit cell adhesion involved in several pathological responses, and methods of producing such derivatives. The invention further relates to pharmaceutical compositions and diagnostic agents containing these derivatives, and methods using these pharmaceutical compositions and diagnostic agents.
Endothelial cells and various circulating cells of the hematolymphoid system express unique surface glycoproteins known as selecting, which mediate intercellular adhesion. (K.-A. Karlsson, TIPS 12: 265-272 (1991)). Intercellular adhesion plays an important role in many pathological responses. For example, the adherence of platelets and leukocytes to vascular surfaces is a critical component of the inflammatory response and is part of a complex series of reactions involving the simultaneous and interrelated activation of the complement, coagulation and immune systems. In addition, leukocyte adherence to vascular endothelium is a key initial step in the migration of leukocytes to tissues in response to microbial invasion.
Compounds that block this initial adhesive interaction are thus expected to have efficacy in the treatment of inflammatory diseases such as rheumatoid arthritis, asthma, and psoriasis. Other indications include adult respiratory distress syndrome, reperfusion injury, ischemia, ulcerative colitis, vasculitis, atherosclerosis, and inflammatory bowel disease. (Boschelli et al., U.S. Pat. No. 5,356,926). Synthetic analogs (mimetics) of carbohydrate ligands that bind specifically to selectins, and thus inhibit selectin-mediated intercellular adhesion, have been implicated as promising therapeutics as anti-inflammatories and anti-coagulants (T. A. Springer, L. A. Lasky, Nature 349: 196-197 (1991); T. Feizi, TIBS 16: 84-86 (1991)).
In addition, carbohydrate ligands are involved in bacterial and viral infections and cancer. Carbohydrate ligands are recognition domains for viruses (J. C. Paulson, The Receptors, Vol. II, P. M. Conn, ed., Academic Press, 131 (1985)), bacteria (Stromberg et al., EMBO J. 9: 2001 (1990)) and toxins (Karlsson et al., Sourcebook of Bacterial Protein Toxins, eds. J. E. Alouf, J. H. Freer, Academic Press, 56: 3537 (1990)). Carbohydrate mimetics are thus expected to have efficacy in the prevention and treatment of bacterial and viral infections and sepsis.
Cancer cells express carbohydrate ligands in patterns different from those in normal cells. Carbohydrate mimetics could be used to generate antibodies that recognize the naturally occurring carbohydrate ligands and thus facilitate the diagnosis of cancer. Because leukocyte adherence to vascular endothelium is also relevant to tumor cell metastasis, synthetic analogs that inhibit selectin-mediated intercellular adhesion are expected to have efficacy in the treatment of metastatic conditions. (S.-i. Hakomori, Cancer Cells, Vol. 3, No. 12 (December 1991)).
With regard to selectin-mediated intercellular adhesion, silylated and fucosylated carbohydrate ligands, specifically sialyl-Lewis-X[.alpha.Neu5Ac(2.fwdarw.3).beta.Gal(1.fwdarw.4)[.alpha.Fuc(1 .fwdarw.3)]-.beta.GlcNAc-OR] and sialyl-Lewis-A [.alpha.Neu5Ac(2.fwdarw.3).beta.Gal(1.fwdarw.3)[.alpha.Fuc(1.fwdarw.4)]-.b eta.GlcNAc-OR] (where R is defined as an aglycone having at least one carbon atom), are particularly important. (Schauer, ed., "Sialic Acids" in Cell Biology Monographs, Vol. 10 (1982); Lowe et al., Cell, 63: 475-485 (1990)). Both chemical (Ratcliff et al., U.S. Pat. No. 5,079,353) and chemical/enzymatic (A. Venot et al., PCT/CA 92/00251) syntheses of these compounds have been developed. These processes, however, are highly complex.
Therefore, research is presently underway to develop analogs that are easier to synthesize, but have equal or greater selectin binding affinity, than the naturally occurring carbohydrate ligands. Toward this end, several analogs, containing particular residue substitutions, have been synthesized. For example, neuraminic acid was replaced by lactic or glycolic acid, fucose was replaced by glycerol or trifluoromethylfucose, and N-acetylglycosamine was replaced by glycosamine or glucose (PCT/US 92/09870). Substitution of neuraminic acid by sulfate or phosphate has likewise been described (PCT/CA 92/00245). In addition, the replacement of glucosamine with a chain of at least 2 carbon atoms has been described (WO 92/18610). These efforts, however, have not elucidated whether oligo- or polyvalent structures can be synthesized from these substituted compounds. Furthermore, no methods have been described for the synthesis of compounds that contain a diol or similar group instead of the anomeric sugar.
To date, an efficient, enzymatic large-scale synthesis has been developed only for native sialyl-Lewis-X and sialyl-Lewis-A having slight modifications (C. H. Wong et al., WO 92/16640 and U.S. Pat. No. 5,162,513). These analogs, however, have the disadvantages of both a low affinity for corresponding selectins and a low in vivo stability (active substances are not orally available).